Epidermis has been an important model tissue for the study of epithelial tumorigenesis. In man, epidermis is one of several stratified squamous epithelial (i.e., cervix, esophagus, pharnyx) that exhibit a high incidence of cancer, and in animal models, topically applied chemicals induce tumors of epidermal origin. The primary goal of this research project is to use a human epithelial cell type (epidermal keratinocytes) to isolate and describe rare carcinogen-induced variants in growth control and terminal differentiation. Valid oncogenic phenotypes in human epithelial cells are a prerequisite for the study of the genetic basis of chemically-induced carcinogenesis in epithelial tissues. The techniques for the selection of carcinogen-induced keratinocyte variants are straightforward and based on cell-type specific growth requirements for normal keratinocytes and naturally occurring squamous cell carcinoma lines. Specifically, transformed phenotypes of carcinogen treatment human keratinocytes will be selected by the following schemes; 1.) clonal growth and serial cultivation in the absence of EGF or cholera toxin, 2.) alterations in the rate of terminal differentiation as measured by the ability to reinitiate growth in surface after suspension in semi-solid medium, and 3.) differences in the ability to form a terminal differentiation end product, cornified envelopes. The tumorigenicity of keratinocytes variants with altered growth factor or differentiation properties will be monitored in nude mice. Carcinogen-induced mutation frequency at the hprt locus will be measured in order to determine the relationship, if any, between mutation frequency and transformation frequency of the chemically treated human epidermal keratinocytes. The ability to detect mutational events in keratinocytes coupled with the ability to identify of cell type specific markers of epithelial transformation provides a human model where the relevance of single locus mutation assays for the prediction of carcinogenicity in humans can be assessed.